Antibodies that inactivate antihemophilic factor (AHF, factor VIII) develop in approximately 10 percent of multiply-transfused patients with severe hemophilia as well as in individuals without a prior hemorrhagic disorder who develop an autoantibody that inactivates factor VIII coagulant activity. This project seeks an understanding of the mechanism by which these antibodies inactivate factor VIII coagulant activity, the reason for the lack of immunoprecipitate formation by these antibodies, and the nature of the antigen determinants which they react. It also seeks information about molecular (immunologic) heterogeneity in normal factor VIII and a better definition of the comparative immunologic properties of normal and hemophilic factor VIII. As a part of these studies, IgG and Fab' will be purified from high-titer human anti-factor VIII sera. Antigenic determinants related to the factor VIII coagulant site will be identified by detection of specific immune complexes with anti-factor VIII. A radioimmunoassay for these determinants will also be developed by specific purification 125-I-labeled human anti-factor VIII. Recent studies have identified both large (greater than 10 to the 6th power daltons) as well as small (less than 300,000 daltons) immune complexes when human anti-factor VIII are incubated with normal cryoprecipitate. We will seek more definitive characterization of the small complexes, of the relationship of factor VIII components in the presence and absence of human anti-factor VIII, and of the binding affinity of human anti-factor III with coagulant site antigens.